1. Field of the Invention
The present invention relates to a quantitative determination method for heparan sulfate contained in body fluid specimens, which method is useful for identifying various diseases. The present invention also provides means for judging the condition of diabetic nephropathy by determination of heparan sulfate contained in urine specimens by use of the quantitative determination method of the present invention. The present invention further provides means for diagnosing hepatic diseases and rheumatoid arthritis by determination of heparan sulfate contained in blood specimens by use of the quantitative determination method of the present invention.
2. Description of the Related Art
Diabetes is known to cause various complications. Among the fatal complications caused by diabetes is "diabetic nephropathy," which results in grave renal failure.
In healthy kidneys, a negative charge in the heparan sulfate sugar chain of heparan sulfate proteoglycan that is present in the glomerular basement membrane functions as a "charge barrier" against protein and the like in blood and prevents protein and the like from being deposited on the glomeruli or leaking into urine (Proc. Natl. Acad. Sci. USA 76:1303, 1979).
However, it often happens that the glomerular basement membrane of a patient suffering from diabetic nephropathy is damaged and the function as a "charge barrier" is lost. For diagnosis of diabetic nephropathy, there has conventionally been employed a method in which albumin that has leaked into urine after passing through the "charge barrier" is measured and the quantity is compared with a normal value of a healthy person, to thereby determine incipiental diabetic nephropathy.
As a method for measuring heparan sulfate, a method of using anti-heparan sulfate antibody JM403 has been known. However, the anti-heparan sulfate antibody is known to exhibit cross-reactivity to hyaluronic acid (Kidney International, 41(1992) pp115-123). Therefore, the method, being unable to provide accurate measurement of heparan sulfate, is impractical as a method for measuring only heparan sulfate, and cannot be used in the diagnosis of diseases.
Patients suffering from diabetes run a high risk of developing diabetic nephropathy. Diabetes and diabetic nephropathy are classified as follows depending on urine albumin: 1) diabetes (high blood sugar level/creatinine-corrected (CR) urine albumin: less than 12 mg/gCR; also described as DM1 hereinafter), 2) incipiental diabetic nephropathy (high blood sugar level/creatinine-corrected (CR) urine albumin: 12 mg/gCR or higher and less than 200 mg/gCR; also described as DM2 hereinafter), and 3) diabetic nephropathy (high blood sugar level/creatinine-corrected (CR) urine albumin: 200 mg/gCR or higher; also described as DM3 hereinafter).
Renal pathosis is not detected in patients in the stage of DM1. Patients in the stage of DM2 have minor renal pathosis, and their condition may be improved to DM1 by proper treatment. However, patients in the stage of DM3 have grave renal pathosis, which may hardly be improved even by proper treatment.
Thus, because prognosis after a true onset of diabetic nephropathy is very unfavorable, the progression of diabetic nephropathy must be diagnosed as early as possible.
However, the level of albumin contained in an urine specimen, which has conventionally been used as a diagnostic index for diabetic nephropathy, varies even in a healthy person, and therefore it is difficult to accurately judge, solely by the albumin level, the condition of patients suffering from diabetes whose stage may be in progress from diabetes (DM1) to incipiental diabetic nephropathy (DM2), or further to diabetic nephropathy (DM3).
In addition, there are many cases in which proper treatment is commenced when, based on a change in the level of albumin contained in a urine specimen, a patient is given a definite diagnosis of incipiental diabetic nephropathy or diabetic nephropathy, yet the treatment turns out to be ineffective, leading to grave nephropathy or further to renal failure.
Accordingly, a first subject of the present invention is the provision of means for specifying as early as possible patients suffering from diabetes who according to the classification based on the level of albumin in a urine specimen do not yet have incipiental diabetic nephropathy, but are in the course of the progression to incipiental diabetic nephropathy, or patients suffering from incipiental diabetic nephropathy who are in the course of the progression to diabetic nephropathy.
In many hepatic diseases, pathological changes occur very slowly in their early stage. Thus, it often happens that by the time pathosis is detected, the condition has progressed too far for the patient to be saved or the patient must undergo treatment for many years.
Conventional methods for detecting hepatic diseases include assays of hepatopathy markers such as blood cholinesterase, hepaplastin, GOT, GPT, .gamma.GTP, bilirubin, and immunoglobulin; as well as a liver function test. Also, measurement of urine bilirubin is used to screen for hepatic diseases. In addition, image diagnosis by CT scanning is used to detect hepatic diseases. Moreover, for diagnosis of the causes of hepatic diseases, anti-hepatitis virus antibody/antigen is tested.
There is also provided a method for diagnosis of hepatic diseases by the detection of fibrogenesis caused by hepatic diseases. For example, a histologic detection method in which a biopsy of the liver is performed by laparoscopy or the like is provided.
However, these diagnostic methods have drawbacks such as sampling errors and invasion of the living body. A proposed method to detect fibrogenesis in the liver caused by hepatic diseases based on the rise in blood .gamma.-globulin or the like has also been found unsatisfactory. Accordingly, none of these methods are effective for detecting hepatic diseases.
The criteria set by the American Rheumatology Society are currently used to diagnose rheumatoid arthritis. A blood test, which is an objective test among the criteria, includes only one measurement item, i.e. rheumatoid factor. However, a test for rheumatoid factor is known to be not highly specific. A test for anti-rheumatoid factor antibody (IgG-RF) is also performed, but used only in patients who have already been diagnosed as having rheumatism, so as to judge the seriousness of their arthritis and whether they have angitis or not.
Thus, an effective method for diagnosis of rheumatoid arthritis has not yet been developed, and the diagnosis is made based on subjective symptoms reported by patients. Although the level of hyaluronic acid in blood is known to rise in patients suffering from rheumatoid arthritis, detecting rheumatoid arthritis based on this finding is difficult and does not provide an accurate method for detection. Another known method of judging the condition of rheumatoid arthritis uses an index based on the degree of the patient's mobility in daily life and ability to exercise such as degrees of dysfunction (class classification by Steinbrocker et al.), activity index (Ransbury index), conditions of joints (stage classification). The method permits judgment of conditions, but does not provide an accurate judgment because the judgment is not quantitative and is based on subjective symptoms reported by patients, which may vary a great deal among patients.
As described above, various means for detecting hepatic diseases and rheumatoid arthritis have been provided, but none of the methods are satisfactory.
As a result, accurate diagnosis of hepatic diseases and rheumatoid arthritis requires a large number of tests, which is complicated and unsuitable for screening a large number of specimens.
Accordingly, a second subject of the present invention is the provision of means for diagnosing hepatic diseases and rheumatoid arthritis accurately and which permits simple primary screening of a large number of specimens.